restriction enzymes/maps and recontaminate DNA, Flashcards
what is a nucleas and that are the 2 kind
enzyme that breaks down nucleic acid
DNase, and RNase
DNase exonuclease
chews up DNA from the ends
DNase endonuclease
cuts DNA in the middle of the molecule can be non specific or specific
restriction enzymes are
endonuclease that cut DNA at specific sequence
EcoR1 recognizes _______ and leaves a _bp sticky end
“GAATTC” , 6
what enzymes leave a blunt end
Alu1,Haelll,Sma1
Alu recognizing _ bp, and Not1 recognizing _ bp
4,8
if there is 5 sites of a GAATC that EcoR1 cuts how many peices will be cut
5 pieces
why is DNA negatively charges
the phosphate groups PO4-
in gel electrophoresis
short dna molecules will move faster through the gel than large strands
what is dna in electrolysis stained with
ethiduim bromide
what is recombination dna
DNA that has been formed artificially by combining constituents from different organisms.
what are 3 things needed to make recombinate dna
purified DNA, vector, dna ligase
what are useful properties of restriction enzymes
cut DNA at specific sites, can produce sticky ends
what is a cloning vector
small DNA molecule that can replicate in a host bacterial cell (must have a origin or replication)
what are some examples of cloning vectors
plasmid or phage chrosome
what should a suitable cloning vector have
1) should carry a selectable genetic markers
2) have one or more unique restriction sites - squenece that is cute by a restr. enzyme only once in the vector
3) should be a way to easily distinguish recombinate DNA
what are the 3 possiblites when cut DNA and vectors with sticky ends are mixed
DNA fragment cirlce forms
the vector cirles re-forms
recombinant DNA forms
add ____ to the recombin. mixture to covalently join ends
ligase
what help you determine whether the bacteria took up vector circle or recombinat DNA
the bacteria is lacZ negative and the plasmid contains the lacZ gene with the coding region for beta (B-galactosidase)
how does X-gal work
when colonies are grown in X-gal the ones that produce B-galactosidase will turn blue which means that it is the vector DNA, the recomb. Dna will be white
what is genomic DNA
all of an organisms genes inclung introns and junk da
genomic library
collection of 100’s of deff. recomb. DNA molecules made by ligating genomic DNA fragments into cloning vector
cDNA library
is used for protein coding and is only comprised or coding regions
